Early-in-life exposure to painful and/or inflammatory stimuli can produce permanent changes in the neuroanatomical and neurophysiological substrates that process nociceptive stimuli in both somatic and gastrointestinal systems. We propose that early-in-life bladder inflammation also may predispose an individual to experience increased bladder sensitivity as an adult. As part of an NIDDK-funded R21 grant, preliminary studies showed that neonatal, but not adolescent, exposure to bladder inflammation led to a hypersensitive nociceptive and reflex state in female rats raised to adulthood, as manifested by increased visceromotor reflexes (VMRs) and arterial blood pressure (ABP) responses to phasic urinary bladder distention (UBD), decreased threshold for micturition reflexes in cystometrographic (CMC) analyses, and increased baseline frequency of micturition. These outcomes also are consistent with two primary symptoms of the painful bladder syndrome interstitial cystitis (IC): enhanced sensory (pain-urgency) and reflex responses (i.e., reduced bladder capacity) to bladder distention. These data served to generate the overall hypothesis of the present proposal. Specifically, we propose the following: An alteration in neuronal sensory substrates is initiated by early-in-life bladder inflammation. This process enhances susceptibility to the development of a hypersensitive state as an adult, especially when a second bladder insult occurs, and is manifested by lowered intravesical pressure thresholds for micturition reflexes and enhanced bladder-related nociceptive responses. This results in urinary dysfunction and pathological urinary bladder pain as an adult. The general hypothesis is tested in three specific aims that determine in a quantitative fashion the effect of neonatal, and in some cases adolescent, bladder inflammation on: (1) VMRs and c-fos expression to phasic UBD, baseline micturition frequency, and micturition reflexes in CMG tests of adult rats, (2) the structure of the bladder, and the density and spinal distribution of bladder afferents in studies of histopathology using protein gene product 9.5 (PGP 9.5), wheat germ agglutinin horse-radish peroxidase (WGA-HRP), substance P, CGRP, and TRPV1 immunhistochemisty, and (3) responses of C- and A-5 bladder afferents to phasic UBD in adult rats. We believe these systematic studies, based on an innovative hypothesis, will lay the groundwork for potential novel therapeutic modalities for the treatment of urinary bladder pain by identifying the substrates for the development of bladder hypersensitivity. Translation to the treatment of painful bladder syndromes like 1C would be highly probable.